建立高效液相層析法及液相層析電灑游離串聯質譜法分析穀類中真菌毒素Zearalenone   全文下載

Determination of Zearalenone in Cereals by High-Performance Liquid Chromatography and Liquid Chromatography–Electrospray Tandem Mass Spectrometry

Zearalenone（玉米赤黴毒素）是一種由鐮刀菌所產生之真菌毒素，常見於食品及飼料中。本研究利用免疫親合管柱淨化穀類樣品，續以高效液相層析（HPLC）及液相層析電灑游離串聯質譜（LC/ESI-MS-MS）建立快速、準確的分析方法。先以90% 乙?溶液萃取穀類製品中之玉米赤黴毒素，再經ZearalaTestTM 免疫親合管柱淨化後得到萃取液。在HPLC 之分析方面，螢光檢測激發波長274 nm、發射波長440 nm，移動相為50% 乙?溶液，流速1 mL/min。在回收率測試部分，於穀類檢體中添加5、20、100 及200 ppb 之玉米赤黴毒素，平均回收率介於66.4-96.1%，方法偵測極限為5 ppb。在LC/ESI-MS-MS 之分析中，採用負離子電灑游離法，以選擇反應偵測模式檢測，選擇m/z 317 為母離子、m/z 175 為子離子。方法偵測極限為3 ppb。市售穀類製品之調查結果，在7 件玉米製品中檢出4 件含有玉米赤黴毒素，惟含量皆低，介於7.9~9.0 ppb 之間。另在7 件綜合穀類製品、5 件小麥製品、5 件白米製品及2 件燕麥製品中皆未檢出。以本次檢出玉米赤黴毒素之最高含量推算國人可能攝食量之結果，遠低於國際間規定之每日攝取容許量。

Zearalenone is a mycotoxin produced mainly by fungi belonging to the genus Fusarium in foods and feeds. A rapid and accurate method to quantify zearalenone in cereals was described. The determination of zearalenone was performed using immunoaffinity column for clean-up, high-performance liquid chromatography (HPLC)/fluorescence for quantification and liquid chromatography/ electrospray tandem mass spectrometry (LC/ESI-MS-MS) for confirmation. Cereal samples were extracted with acetonitrile- water (90:10, v/v) prior to a Vicam ZearalaTestTM immunoaffinity column clean-up. Zearalenone was eluted from the column with methanol and quantified by HPLC with fluorescence detection (λex = 274 nm, λem = 440 nm). A mobile phase of acetonitrilewater (50:50, v/v) and a flow rate of 1.0 mL/min-1 were used. Average recoveries of zearalenone from cereals spiked at levels of 5, 20, 100 and 200 ppb ranged from 66.4% to 96.1%. The limit of quantitation was 5 ppb. The selective determination of zearalenone was achieved by LC/MS-MS with an electrospray (ESI) ionization interface. Using the negative ion mode, the parent ion of m/z 317 and the product ion of m/z 175 were selected. The quantification limit of 3 ppb was achieved. Seven mixed cereals samples, seven corn samples, five wheat samples, five rice samples and two oat samples were analyzed. The results showed that zearalenone was detected in four corn samples ranging from 7.9 to 9.0 ppb. Using the consumption data from “Nutrition and Health Survey in Taiwan 1993~1996”, the zearalenone consumption for Taiwanese adults (male and female are 0.00297 and 0.00478 μg/kg b.w. respectively) was much lower than the tolerable daily intake of 0.5 μg/kg b.w. established by JECFA.